全文获取类型
收费全文 | 81576篇 |
免费 | 19128篇 |
国内免费 | 24篇 |
出版年
2023年 | 182篇 |
2022年 | 450篇 |
2021年 | 1412篇 |
2020年 | 2670篇 |
2019年 | 4306篇 |
2018年 | 4738篇 |
2017年 | 4900篇 |
2016年 | 5537篇 |
2015年 | 6398篇 |
2014年 | 6498篇 |
2013年 | 7289篇 |
2012年 | 6246篇 |
2011年 | 5736篇 |
2010年 | 5618篇 |
2009年 | 4190篇 |
2008年 | 4146篇 |
2007年 | 3591篇 |
2006年 | 3211篇 |
2005年 | 3048篇 |
2004年 | 2864篇 |
2003年 | 2487篇 |
2002年 | 2224篇 |
2001年 | 1843篇 |
2000年 | 1677篇 |
1999年 | 1336篇 |
1998年 | 534篇 |
1997年 | 462篇 |
1996年 | 352篇 |
1995年 | 319篇 |
1994年 | 326篇 |
1993年 | 284篇 |
1992年 | 511篇 |
1991年 | 490篇 |
1990年 | 438篇 |
1989年 | 407篇 |
1988年 | 340篇 |
1987年 | 323篇 |
1986年 | 287篇 |
1985年 | 243篇 |
1984年 | 196篇 |
1983年 | 180篇 |
1982年 | 157篇 |
1981年 | 152篇 |
1980年 | 144篇 |
1979年 | 175篇 |
1978年 | 161篇 |
1976年 | 131篇 |
1975年 | 138篇 |
1974年 | 156篇 |
1973年 | 127篇 |
排序方式: 共有10000条查询结果,搜索用时 15 毫秒
991.
Summary A simple experimental method is devised to determine the fraction of plasmid-harboring cells in a bioprocess employing recombinant mammalian cells. The fraction of plasmid-harboring cells decreased as serum content in the growth medium decreased. The relatively higher increase in the generation time of the plasmid-harboring cell was primarily responsible for this decrease. The mathematical expression obtained for this fraction in terms of the two parameters, i.e. the generation time ratio and the plasmid-loss probability, could represent the experimental data extremely well. The numerical values of these parameters could show the inherent insight of the system. It was found that the data plot against time can draw us to a misleading conclusion of the absence of the effect of serum concentration. 相似文献
992.
Summary The cloning of glucoamylase geneSTA using theSUC2 promoter intoSaccharomyces cerevisiae was performed. The signal sequence ofSTA gene was used for the secretion of glucoamylase protein. The plasmid constructed in this way was named YEpSUCSTA and its expression was identified. The expression of YEpSUCSTA was repressed in the presence of glucose in growth medium, but derepressed when glucose became depleted. YEpSUCSTA showed the similar efficiency of glucoamylase secretion as YEpSTA-F which has the entireSTA gene. Glucoamylase activity in starch-glucose medium was largely increased because cell mass and plasmid stability were high in biosynthesis phase compared to extracellular glucoamylase activities in media which starch or glucose was the only carbon source. 相似文献
993.
Summary Clinical dextran with desired molecular weight was produced continuously in the two-stage reactor. Cells ofLeuconostoc meseteroides B512F cultivated in the first reactor were transferred to the second reactor where sucrose and primer were added for clinical dextran production. By using this two-stage reactor, the fraction of desired clinical dextran increased significantly when observed with gel permeation chromatography. 相似文献
994.
995.
Manfred Braun Jong Min Kim Rolf D. Schmid 《Applied microbiology and biotechnology》1992,37(5):594-598
Summary The soil isolate Cellulomonas cellulans AM8 produces an extracellular l-amino acid oxidase (L-AAO) with broad substrate specificity. The strain produced up to 0.35 unit (U)/ml of the extracellular L-AAO in a simple medium containing glycerol and yeast extract. The enzyme was easily purified up to 30 U/mg protein using Phenyl-Sepharose fast flow. The purified enzyme migrated as single band on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) with a molecular mass of 55 kDa. On native PAGE the molecular mass was approx. 300 000 kDa, which may be due to aggregation. With the exception of glycine, proline, and threonine, all the amino acids normally constituting proteins were oxidized. The V
max values from 0.7 to 35.2 U/mg for aspartic acid and lysine, respectively, and the K
m values from 0.007 to 7.1 mm for cysteine and valine, respectively, were obtained at 25° C and pH 7.0 in oxygen-saturated solutions. The L-AAO had a pH optimum of 6.5–7.5. It was stable for several months at — 30° C and for some days at 35° C. Ferricyanide served as an electron acceptor with a V
max of 50 U/mg and K
m for 0.3 mm with phenylalanine as the substrate.
Correspondence to: R. D. Schmid 相似文献
996.
We show that bacteriophage lambda DNA fragments microinjected into the macronucleus of the ciliated protozoan Paramecium can replicate as unit-length linear molecules. These linear DNA molecules are substrates for the addition of Paramecium telomeres by an endogenous telomerase. The linear DNA pieces can exist at copy numbers much higher than that of typical endogenous macronuclear chromosomes. We show that the copy number of injected DNA many fissions after microinjection reflects that of the original input copy number, suggesting that active control of copy number does not occur. Instead, the results suggest that injected DNA is replicated once per cell division. 相似文献
997.
998.
An intracellular form of phospholipase A2 was purified about 47,500-fold to near homogeneity from bovine platelets 100,000 x g supernatant by sequential use of column chromatographies on Heparin-Sepharose, DEAE-Sephacel, Butyl-Toyopearl, Sephacryl S-300, DEAE-5PW HPLC, TSK G 3000 SW HPLC and Mono Q FPLC. The final preparation showed a single band on SDS-polyacrylamide gel, and its molecular mass was estimated to be approximately 100,000 daltons. The purified PLA2 showed maximal activity at alkaline pH(pH 9.0-10.0) and considerable activity at 0.3-1.0 microM calcium concentration. It hydrolyzed phosphatidylcholine containing arachidonate at sn-2 position with high selectivity in comparison to linoleate. 相似文献
999.
Rat brain plasma membranes were solubilized in detergent and a glycoprotein-enriched fraction was obtained by lectin affinity chromatography. This glycoprotein fraction contained insulin receptors, as well as protein kinases capable of phosphorylating some exogenously added substrates such as MAP2 (microtubule associated protein 2) and MBP (myelin basic protein), but not ribosomal protein S6. Phosphoamino acid analysis of MAP2 and MBP showed that phosphotyrosine residues, as well as phosphoserine/phosphotheronine residues, were present in both proteins under basal conditions. Whereas the addition of insulin to the rat brain membrane glycoprotein fraction in vitro had no effect on MAP2 phosphorylation, MBP phosphorylation was stimulated 2.7-fold in response to insulin. This phenomenon was dose-dependent, with half-maximal stimulation of MBP phosphorylation observed with 2 nM insulin. Phosphoamino acid analysis of MBP indicated that insulin stimulated the phosphorylation of tyrosine residues nearly three-fold, whereas the phosphorylation of serine or threonine residues was not increased. These results identify MBP as a substrate for the rat brain insulin receptor tyrosine-specific protein kinase in vitro. 相似文献
1000.
An endonuclease activity in human polymorphonuclear neutrophils that removes 8-hydroxyguanine residues from DNA+ 总被引:9,自引:0,他引:9
M H Chung H S Kim E Ohtsuka H Kasai F Yamamoto S Nishimura 《Biochemical and biophysical research communications》1991,178(3):1472-1478
An endonuclease that specifically removes 8-hydroxyguanine (oh8Gua) from DNA has been isolated from Escherichia coli. As the amount of oh8Gua produced in DNA of X-ray-irradiated mice is known to decrease with time after irradiation, an attempt was made to find a similar activity in human polymorphonuclear neutrophils (PMNs) using a synthetic dsDNA containing oh8Gua as a substrate. The PMN enzyme was isolated free of other DNases, and found to cleave the substrate DNA simultaneously at 2 sites, the phosphodiester bonds 5' and 3' to oh8Gua, producing free hydroxyl and phosphate groups, respectively. The enzyme showed almost no activity on DNAs containing other kinds of modified base tested or mismatched DNA. Thus human cells also contain an endonuclease that specifically removes oh8Gua residues from DNA. 相似文献